Proteinuria is the hallmark and surrogate marker of many primary and secondary glomerular diseases resulting in serious kidney malfunctions that occur in patients with diabetes and hypertension. To enhance the diagnostic and therapeutic needs related to this serious disease, this project seeks to quantify glomerular permeability in vivo with unmatched precision by: (1) applying a new generation of semiconductor nanocrystals (CdSe/ZnS quantum dots) as highly accurate size rulers and very sensitive electrostatic nanosensors combined with (2) a recently developed quantitative two-photon excitation microscopic imaging technique that uses ultrafast near-infrared laser for deep tissue penetration. Currently, we are able to image individual glomeruli and peritubular structures inside a live animal kidney. We hypothesize that the use of these novel quantum dot (QD) systems in conjunction with in vivo two-photon microscopy will increase vastly the accuracy and sensitivity of our imaging by outperforming traditional dextran probes used in kidney studies, resulting in a better understanding of glomerular disease processes and leading to the development of early clinical tests to quantify glomerular permeability. We predict this outcome because quantum dots are rigid and spherical in shape, and can be produced with high monodispersity, high photostability and brightness as well as adjustable surface chemical properties. A "size ruler"-like accuracy can be achieved by applying thin bioinert polar coatings, whereas neutral-polar (Ser-capped) and negatively charged (Asp-capped) peptide-coated QDs will be used to create protein-like surface properties. The goal of this interdisciplinary project is to combine expertise concerning the use of cutting-edge nanoparticle probes for biomedical applications (Naumann, Long) and the state-of-the-art optical imaging (Yu, Molitoris) to develop a significantly improved and enabling imaging/probe system for intravital kidney imaging. The proposed study will apply this new imaging strategy, highly relevant to a major theme outlined in the NIH Roadmap in areas of Quantitative Imaging and Nanomedicine, to provide a basic understanding of the functions of glomerular filtration barrier and insight into the correlation between glomerular damage and the onset of proteinuric symptoms. The results of our study will offer valuable information important for early diagnosis, improving the quality of patient care, and finding a future cure.